title: Uracil excision repair in Mycobacterium tuberculosis cell-free extracts.
abstract: Uracil excision repair is ubiquitous in all domains of life and initiated by uracil DNA glycosylases (UDGs) which excise the promutagenic base, uracil, from DNA to leave behind an abasic site (AP-site). Repair of the resulting AP-sites requires an AP-endonuclease, a DNA polymerase, and a DNA ligase whose combined activities result in either short-patch or long-patch repair. Mycobacterium tuberculosis, the causative agent of tuberculosis, has an increased risk of accumulating uracils because of its G + C-rich genome, and its niche inside host macrophages where it is exposed to reactive nitrogen and oxygen species, two major causes of cytosine deamination (to uracil) in DNA. In vitro assays to study DNA repair in this important human pathogen are limited. To study uracil excision repair in mycobacteria, we have established assay conditions using cell-free extracts of M. tuberculosis and M. smegmatis (a fast-growing mycobacterium) and oligomer or plasmid DNA substrates. We show that in mycobacteria, uracil excision repair is completed primarily via long-patch repair. In addition, we show that M. tuberculosis UdgB, a newly characterized family 5 UDG, substitutes for the highly conserved family 1 UDG, Ung, thereby suggesting that UdgB might function as backup enzyme for uracil excision repair in mycobacteria.
authors.affiliation: Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Pradeep
authors.initials: P
authors.lastname: Kumar
authors.forename: Sanjay Kumar
authors.initials: SK
authors.lastname: Bharti
authors.forename: Umesh
authors.initials: U
authors.lastname: Varshney
mesh_terms: D001426:Bacterial Proteins; D004249:DNA Damage; D004260:DNA Repair / Q000235:genetics*; D006801:Humans; D020102:Mycobacterium smegmatis / Q000235:genetics* / Q000378:metabolism; D009169:Mycobacterium tuberculosis / Q000235:genetics* / Q000378:metabolism; D014498:Uracil / Q000378:metabolism; D051981:Uracil-DNA Glycosidase / Q000235:genetics* / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D014498:Uracil; D051981:Uracil-DNA Glycosidase